How does the Cas9 nuclease locate a specific 20‐nucleotide target sequence in a crowded intracellular environment packed with mega bases of distracting non‐target DNA? Previously, it was shown that Cas9 finds DNA targets via three‐dimensional diffusion. In this issue of The EMBO Journal, Globyte et al (2019) reveal another dimension of the search process, which involves short‐range one‐dimensional sliding. These results have implications for understanding the natural function of Cas9 and its applications in genome engineering experiments.

See also: V Globyte et al

The EMBO Journal (2019) e101474

Cas9 is an RNA‐guided nuclease that is routinely used for targeted genome engineering in eukaryotic cells. However, the roles and responsibilities of genome engineers are profoundly different from the natural selective pressures that optimized Cas9 for protection from invading genetic parasites, like phages. To optimize Cas9 for applications in genome engineering, it is critical to understand the biophysical mechanisms that govern target identification. In this issue of The EMBO Journal, Globyte et al (2019) show that the Cas9 target search process involves a combination of 1D sliding and 3D diffusion.

Unwinding all the double‐stranded DNA (dsDNA) in a cell to find a complementary target would be a slow and energetically expensive process. Instead, target identification by Cas9 first relies on detection of a …

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